The creation of nanostructure is profound for the generation of nanobiosensors in several medical diagnosis. Here, we employed an aqueous hydrothermal route using Zinc-oxide (ZnO) and Gold (Au), which under optimal conditions formed an ultra-crystalline rose-like nanostructure textured with nanowires on the surface, coined as "spiked nanorosette." The spiked nanorosette structures was further characterized to possess crystallites of ZnO and Au grains with average sizes of 27.60 and 32.33 nm, respectively. The intensity for both ZnO (002) and Au (111) planes of the nanocomposite was inferred to be controlled by fine-tuning the percentage of Au nanoparticles doped in the ZnO/Au matrix, as referred by X-ray diffraction analysis. The formation of ZnO/Au-hybrid nanorosettes were additionally verified by the distinct corresponding peaks from photoluminescence and X-ray photoelectron spectroscopy, supported by electrical validations. The biorecognition properties of the spiked nanorosettes were also examined using custom targeted and non-target DNA sequences. The DNA targeting capabilities of the nanostructures were analyzed by Fourier Transform Infrared and electrochemical impedance spectroscopy. The fabricated nanowire-embedded nanorosette exhibited a detection limit at the lower picomolar range of 1 × 10-12 M, with high selectivity, stability and reproducibility and good linearity, under optimal conditions. Impedance-based techniques are more sensitive to the detection of nucleic acid molecule whereas this novel spiked nanorosette demonstrate promising attributes as excellent nanostructures for nanobiosensor developments and their potential future application for nucleic-acids or disease diagnostics.
Clustered regularly interspaced short palindromic repeats (CRISPR) have established itself as a frontier technology in genetic engineering. Researchers have successfully used the CRISPR/Cas system as precise gene editing tools and have further expanded their scope beyond both imaging and diagnostic applications. The most prominent utility of CRISPR is its capacity for gene therapy, serving as the contemporary, disease-modifying drug at the genetic level of human medical disorders. Correcting these diseases using CRISPR-based gene editing has developed to the extent of preclinical trials and possible patient treatments. A major impediment in actualizing this is the complications associated with in vivo delivery of the CRISPR/Cas complex. Currently, only the viral vectors (e.g., lentivirus) and non-viral encapsulation (e.g., lipid particles, polymer-based, and gold nanoparticles) techniques have been extensively reviewed, neglecting the efficiency of direct delivery. However, the direct delivery of CRISPR/Cas for in vivo gene editing therapies is an intricate process with numerous drawbacks. Hence, this paper discusses in detail both the need and the strategies that can potentially improve the direct delivery aspects of CRISPR/Cas biomolecules for gene therapy of human diseases. Here, we focus on enhancing the molecular and functional features of the CRISPR/Cas system for targeted in vivo delivery such as on-site localization, internalization, reduced immunogenicity, and better in vivo stability. We additionally emphasize the CRISPR/Cas complex as a multifaceted, biomolecular vehicle for co-delivery with therapeutic agents in targeted disease treatments. The delivery formats of efficient CRISPR/Cas systems for human gene editing are also briefly elaborated.
Leptospirosis is a potentially life-threatening zoonosis caused by pathogenic Leptospira and for rapid diagnostics, direct detection is desirable. LipL32 protein is the most suitable biomarker for direct detection. DNA aptamers are sought to be generated against LipL32 by Systemic Evolution of Ligands via Exponential Enrichment (SELEX). LepDapt-5a is the most potent aptamer candidate among all the candidates, as determined by direct Enzyme-linked Aptasorbent Assay (ELASA). LepDapt-5a was predicted to form a G-quadruplex structure as predicted by QGRS Mapper and validated experimentally by direct ELASA. The diagnostic potential of the aptamer was further tested on a direct and sandwich ELASA platform. A LOD of 106 mL-1 and 105 mL-1 were estimated by direct and sandwich ELASA platforms, respectively, which are within the range associated with leptospiremia levels. The dot blot assay developed was able to attain a LOD of 104 CFU mL-1 against pathogenic Leptospira, which is also within the leptospiremia level. This is the first-ever DNA aptamer and hybrid-heterodimeric aptamer constructed against LipL32. The diagnostic potentiality of the LepDapt-5a DNA aptamer was proven on three major diagnostic platforms, which are direct ELASA, sandwich ELASA, and aptamer-based dot assay.
The present study evaluates the corrosion behavior of poly[xylitol-(1,12-dodecanedioate)](PXDD)-HA coated porous iron (PXDD140/HA-Fe) and its cell-material interaction aimed for temporary bone scaffold applications. The physicochemical analyses show that the addition of 20 wt.% HA into the PXDD polymers leads to a higher crystallinity and lower surface roughness. The corrosion assessments of the PXDD140/HA-Fe evaluated by electrochemical methods and surface chemistry analysis indicate that HA decelerates Fe corrosion due to a lower hydrolysis rate following lower PXDD content and being more crystalline. The cell viability and cell death mode evaluations of the PXDD140/HA-Fe exhibit favorable biocompatibility as compared to bare Fe and PXDD-Fe scaffolds owing to HA's bioactive properties. Thus, the PXDD140/HA-Fe scaffolds possess the potential to be used as a biodegradable bone implant.